Investigative Genetics
Permanent URI for this collectionhttps://hdl.handle.net/20.500.12503/21715
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Browsing Investigative Genetics by Author "Thompson, Lindsey"
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Item Massively parallel sequencing of 68 insertion/deletion markers identifies novel sequence variation for utility in human identity testing(2016-03-23) Song, Bing; Thompson, Lindsey; King, Jonathan; Budowle, Bruce; LaRue, Bobby; Wendt, FrankShort tandem repeat (STR) loci are traditionally used by the forensic science community for kinship, missing persons, and human identity testing. These markers hold considerable value due to their size, ability to be multiplexed, and highly polymorphic nature. However, they are unable to provide phenotypic and biogeographic ancestry estimates and are too large for use in analysis of DNA from highly compromised substrates such as explosives or human remains. Small bi-allelic polymorphisms, such as insertions/deletions (INDELs), have been of considerable interest within the forensic science community for their utility in filling such gaps. These markers range in size from 2-6 base pairs, making them ideal for highly compromised sample types. Additionally, the ease of multiplexing large INDEL panels allows for comparable discrimination power when compared to STRs. Capillary electrophoresis is a current mainstay in the forensic DNA workflow, generating fluorescent signals to detect alleles separated by size. This method is limited by number of dyes simultaneously utilized, number of loci capable of multiplexing, sample throughput, and required amplicon size. Massively parallel sequencing (MPS) provides a solution to these limitations by targeting many loci across the genomes of multiple samples simultaneously with relatively high sequence coverage. Herein, we describe the utility of MPS, using the Nextera™ Rapid Capture Custom Enrichment Kit (Illumina, Inc., San Diego, CA), to sequence 68 INDELs in four major US population groups on the Illumina MiSeq™. We also define a novel application of the STR Allele Identification Tool: Razor (STRait Razor) to analyze INDEL sequences and capture adjacent sequence variation in the form of single nucleotide polymorphisms (SNPs). This application has enabled the discovery of unique allelic variants, which increase the discrimination power and decrease the single-locus and combined random match probabilities of four well-characterized INDELs. These findings suggest that more valuable INDELs for human identification may exist elsewhere in the genome. As such, it is recommended that these four markers be included in future INDEL multiplex panels for human identification due to their enhanced individualization potential.Item Multiplex of INDELs for Human Identification Markers(2016-03-23) Sturm, Sarah; Thompson, Lindsey; Wiley, Rachel; King, Jonathan; LaRue, Bobby; Sage, KellyForensic laboratories commonly use short tandem repeat (STR) loci when comparing an evidentiary profile to that of a reference profile. In commercially available STR kits, the amplified products tend to range from 100- 500 base pairs (bp). For genomic DNA of degraded biological samples, the fragments are usually broken into product sizes of 180-200bps or less. Therefore, degraded biological samples may not produce a full STR profile. Another viable option has been proposed to enable successful typing of some degraded DNA samples. Insertion/ deletion (INDEL) polymorphisms are intergenic regions of the genome in which amplified products can be smaller in length than most STRs. Since forensic genotyping relies on comparison of an evidentiary sample DNA profile with that of a reference sample DNA profile, which usually come from suspects or victims, using highly discriminating markers is desirable. A multiplex panel of human identification (HID) INDEL markers that can individualize people would be beneficial. This project will test the hypothesis that INDELs, which can be used to identify individuals with high discriminatory power, can be developed as a multiplex PCR approach. In testing this hypothesis, primers were designed and multiplexed together to amplify specific INDELs that have been previously identified to be suitable for human identity testing purposes.